Methanotrophs Contribute to Nitrogen Fixation in Emergent Macrophytes

Abstract

Root-associated aerobic methanotroph plays an important role in reducing methane emissions from wetlands. In this study, we examined the activity of methane-dependent nitrogen fixation and active nitrogen-fixing bacterial communities on the roots of <i>Typha angustifolia</i> and <i>Scirpus triqueter</i> using a ¹⁵N-N₂ feeding experiment and a cDNA-based clone library sequence of the <i>nifH</i> gene, respectively. A ¹⁵N-N₂ feeding experiment showed that the N₂ fixation rate of <i>S. triqueter</i> (1.74 μmol h^-1 g^-1 dry weight) was significantly higther than that of <i>T. angustifolia</i> (0.48 μmol h^-1 g^-1 dry weight). The presence of CH₄ significantly increased the incorporation of ¹⁵N-labeled N₂ into the roots of both plants, and the rate of CH₄-dependent N₂ fixation of <i>S. triqueter</i> (5.6 μmol h^-1 g^-1 dry weight) was fivefold higher than that of <i>T. angustifolia</i> (0.94 μmol h^-1 g^-1 dry weight). The active root-associated diazotrophic communities differed between the plant species. Diazotrophic <i>Methylosinus</i> of the <i>Methylocystaceae</i> was dominant in <i>S. triqueter</i>, while <i>Rhizobium</i> of the <i>Rhizobiaceae</i> was dominant in <i>T. angustifolia</i>. However, there were no significant differences in the copy numbers of <i>nifH</i> between plant species. These results suggest that N₂ fixation was enhanced by the oxidation of CH₄ in the roots of macrophytes grown in natural wetlands and that root-associated <i>Methylocystacea</i>, including <i>Methylosinus</i>, contribute to CH₄ oxidation-dependent N₂ fixation.